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Enzymes
Enzymes
Enzyme- highly specific protein catalysts
Enzyme Specificity
Enzyme Specificity
• Lock and Key model
• Induced fit model– “polyaffinity mechanism”- three point attachment
Catalyst
Catalyst- speeds up reaction without being consumed (no effect on equilibrium)
do so by lowering the activation energy of the rxn
activation energy- the amount of energy required to reach the transition state
Catalyst
Catalyst- speeds up reaction without being consumed
do so by lowering the activation energy of the rxn
activation energy- the amount of energy required to reach the transition state
Classes of Enzymes
Enzyme Commission (E.C.) 4.1.1.32
1. Oxidoreductases
Coenzymes
Coenzyme- organic molecule required by an enzyme to catalyze rxn
Most coenzymes are vitamin derivatives (water sol)
Classes of Enzymes
Enzyme Commission (E.C.) 4.1.1.32
1. Oxidoreductases- lactate dehydrogenase
2. Transferases- glucokinase
3. Hydrolases- chymotrypsin, G6Pase
4. Lyases- fumarase
5. Isomerases- phosphoglucoisomerase
6. Ligases- Acyl CoA synthetase
Classes of Enzymes
1. Oxidoreductases
2. Transferases
3. Hydrolases
4. Lyases
5. Isomerases
6. Ligases
Cofactors and Coenzymes
Cofactor- depends on context– either inorganic atom– or inorganic molecule or coenzyme
Coenzyme- organic molecule required by an enzyme for it’s catalytic activity, usually vitamin or vitamin derivative
Coenzymes
1. Oxidoreductases
NAD+/NADH + H+
NADP+/NADPH + H+
FAD/FADH2
NAD+/NADH
Niacin derivative
recognize structure
used for degradation
diffuses in and out of active site
NADP+/NADPH
Almost identical to NAD+
used for synthesis
diffuses in and out of active site
FAD/FADH2
Riboflavin derivative
used for degradation
Prosthetic group
Coenzymes (table of vitamin, coenz form and function)
2. Transferases
TPP
THF
PLP
lipoic acid
vitamin B12
CoASH
6. Ligases
biotin
Kinetics: Rate of Reaction
Kinetics: Rate of Reaction
Kinetics: Rate of Reaction
Kinetics: Rate of Reaction
Michaelis-Menton Kinetics
Eqn.
Vo
Vmax
Km
[S]
Lineweaver-Burk Transformation
Eqn of transformation
slope and intercepts
Michaelis-Menton Kinetics Substrate Concentration
[substrate]
0.00 0.05 0.10 0.15 0.20 0.25 0.30 0.35
velo
city
0.0
0.5
1.0
1.5
Michaelis-Menton Kinetics Enzyme concentration
mL enzyme
0.0 0.3 0.6 0.9 1.2 1.5 1.8
velo
city
0.0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
1.0
Michaelis-Menton KineticsTemperature
temperature, oC
0 10 20 30 40 50 60 70
velo
city
0.0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
1.0
Michaelis-Menton KineticspH
pH
0 2 4 6 8 10 12
velo
city
0
5
10
15
20
25
30
35
40PepsinG6Pase
Michaelis-Menton Kinetics Inhibitors or activators
• Activators- not discussed at this time
• Inhibitors- 3 types
Enzyme Inhibition
Competitive inhibition-
Noncompetitive inhibition-
Uncompetitive inhibition-
Example Problem [S],µmol vo, µmol/min
0.1 0.27
2.0 5
10.0 20
20.0 40
40.0 64
60.0 80
100.0 100
200.0 120
1000.0 150
2000.0 155
Michaelis-Menton Plot
Substrate, µmol
0 500 1000 1500 2000
Vo,
µm
ol/m
in
0
20
40
60
80
100
120
140
160
Example Problem
[S],µmol 1/[S], µmol-1 vo, µmol/min 1/vo, min/µmol
0.1 10 0.27 3.70
2.0 0.5 5 0.2
10.0 0.1 20 0.05
20.0 0.05 40 0.025
40.0 0.025 64 0.0156
60.0 0.0167 80 0.0125
100.0 0.01 100 0.01
200.0 0.005 120 0.0083
1000.0 0.001 150 0.0067
2000.0 0.0005 155 0.0065
Lineweaver-Burk Plot
1/Substrate, 1/µmol
0.0 0.0 0.2 0.3 0.4 0.5
1/V
o, m
in/µ
mol
0.00
0.05
0.10
0.15
0.20
Type of Inhibition
1/[S], mM-1
-1 0 1
v o,
µm
ol/m
L•s
ec
0.00
0.02
0.04
0.06
0.08
0.10
0.12
0.14
0.16
0.18
0.20
-I +I
Enzyme Active Sites
Active site- that region of the enzyme where substrate binds and is converted to product
why the enzyme has to be bigger than substrate
Ways in Which an Enzyme Performs Catalysis
Increase the effective concentration
Stabilize transition state
Put a strain on susceptible bonds
Hold reactants near each other and in the proper orientation
Form covalent bonds with substrate that result in destabilization of substrate
Act as proton donors and acceptors
Nucleophilic/Electrophilic attacks
Amino Acids of the Active Siteget good example of each
X-ray crystallography
mutagenesis
amino acid modifying reagents
Enzyme Regulation
On vs. off
1. Isoenzymes2. Covalent Modification3. Allosterism4. Repression5. Proenzymes
Isoenzymes
LDH example
muscle vs. heart
tetramer
preferential substrate affinity
why?
Covalent Modification
Phosphorylation most common
Others: sulfation, acetylation, methylation
glycogen phosphorylase vs. glycogen synthase
Allosteric Activation/Inhibition
“Other site”
Sigmoidal kinetics
homo- vs. heterotropic
feedback inhibition vs. feedforward stimulation
Repression
molecular biology section
Proenzymes
AKA zymogens
alters the concentration of active enzyme
particularly common with:digestive enzymespeptide hormonesclotting factors
proteolysis is selective
dibasic example
Proinsulin
Other Cleavages
